Isolation and identification of the etiological agent:
Isolation of the etiological agent is attempted in chicken embryos, cell cultures, laboratory animals, culture media or the host species. Cell cultures are most ideal, economic and often give quick results for virus agent. These could be homologous or heterologous; primary, secondary or established cell lines, and of diverse tissue origin (kidney, testes, liver, lung, whole embryo, etc.) depending upon the virus under study. If the viral agent involved is not known, primary kidney or thyroid cell cultures from embryo or young animals of the same (homologous) species should be preferred. The inoculums, after making it free from bacterial and fungal contamination by suitable antibiotic/antifungal treatment, are given 3 to 5 blind passages in a particular cell culture.
Type of cytopathic effects (CPE), demonstration of virus particles/ antigen in the infected cells, haemadsorption, haemagglutination (HA), haemagglutination inhibition (HI), neutralization of CPE with specific serum and physiochemical and biological properties may be studied for identification of the virus. When cell culture facilities are not available or the virus does not grow in any cell culture or produce CPE, chicken embryos may be inoculated in appropriate route i.e. CAM, AC, YS, amniotic cavity or intravenous. Laboratory animals viz. mice, guinea pig, hamster, rabbit, chicken, etc., which support the growth of a number of pathogens may also be used. Route of inoculation for these animals are selected in accordance with the agent being looked for. Strictly host specific viruses those do not grow in any cell culture or chicken embryos, the host species has to be used for its isolation.